This Renewal Application is submitted for the National Center for Research Resources program "Development of High Connectivity Non- Mammalian Models." For some years we have been generating culture techniques and advanced molecular technologies for Strongylocentrotus purpuratus (which have turned out to be useful for other sea urchin species as well). These include methods for egg to egg culture; inbreeding; gene transfer; cis-regulatory analysis; and GFP reporters. Salient successes in the recent grant period include: production of a seven-generation sibling-mated inbred line; characterization and use of single sequence repeat genomic markers; and development of a cis-regulatory element trap (CRE Trap) that works extremely well. We now propose (i) to extend use of the CRE Trap to produce a library of insertions that display interesting regulatory features; (ii) to extend gene transfer technology by new methods to enable better utilization of larval-stage gene expression in the developing adult body plan; (iii) To develop an atlas of rudiment anatomy and study lineage networks in rudiment morphogenesis; (iv) to exploit current observations suggesting that hermaphroditism in S. purpuratus is a heritable trait, and carry out other investigations on genetic aspects of sex in S. purpuratus.